The 42-year-old, who won 14 titles between 2008 and 2012 at Barcelona and took a year off in New York before joining the German side, exposed a deep rift between him and Barça president Sandro Rosell's management team.
"This year there have been too many things that crossed the line," Guardiola told a news conference Thursday at Lake Garda in Italy, where the German team were in pre-season training.
"But using Tito's illness to damage me is something I will never forget," he said in Catalan, denying he had failed to see Vilanova while he was treated in New York for salivary gland cancer.
"I saw Tito in New York and if we did not see each other on other occasions it is because it was not possible, but not on my part." He did not say who exactly had made the accusation.
Barcelona vice president Jordi Cardoner said he was surprised when he heard Guardiola's outburst and he denied the club had made any such claim.
"We have not used Tito's illness to go against him and whoever does such a thing is a bad person," he told Barcelona-based sports daily Mundo Deportivo in an interview published on Friday.
He added he hoped there would be an explanation because he believed Guardiola's relations with himself and Rosell had always been "cordial".
In the same news conference, Guardiola also denied a claim that he had told Brazilian star Neymar's father that Vilanova would not know how to use both Neymar and Lionel Messi in the same squad.
In fact, Guardiola said he had met with Neymar's father and repeatedly told him that his son was doing the right thing by signing with Barcelona.
Vilanova's name was not mentioned in the conversation, he said.
Vilanova had been Guardiola's assistant at Barcelona for five years and he led the Catalan team to their most successful league win in history last season, Guardiola said.
"Do I have to excuse myself for wanting things to go badly for someone who has been a colleague for so long?" he asked. "It is very bad taste and I was not expecting it."
The new Bayern coach said he had asked Barcelona's management to leave him in peace when he left the club.
"The only thing I asked Rosell was to leave me in peace and they have not left me in peace even from 6,000 kilometres away," Guardiola said. "They have not done it, they have not lived up to their word."
Guardiola's quotes were splashed across the pages of Spain's sporting press.
"Guardiola dynamites Barça, " ran the front page of leading sports daily As.
"Guardiola on the attack!" said Mundo Deportivo.